Retinal circuits for local synaptic processing In this application, three widely-known laboratories with complementary expertise that specialize in studying function of retinal circuitry propose to investigate mechanisms of signal processing within On- and Off-bipolar cell types in the mammalian retina. Bipolar cells are essential for the retina because they transmit signals from photoreceptors in the outer retina to amacrine and ganglion cells in the inner retina. They are thought to be non-spiking neurons that relay signals via graded depolarization. However, recently bipolar cells in several species have been reported to generate spike-like calcium transients that may generate bursts of neurotransmitter release similar to those found at most central synapses. In preliminary studies, we showed that mouse bipolar cells also display such spike-like transient events and that independent calcium events can be generated in different axon terminal branches. Using a combination of physiological and computational methods, this project will determine whether the calcium transients in bipolar cell axon terminals represent full- blown spikes, and what computational functions they can perform. Using a live ex vivo mouse retina, we will record responses of bipolar cells and ganglion cells to light stimuli, and construct computational models of the responses to determine the biophysical mechanisms present. The project comprises three specific aims. Aim 1 will determine whether individual bipolar cell axon terminal branches can respond to light independently. This aim will use two-photon calcium imaging in whole-mounted retina to test several hypotheses about the signals that are generated and relayed by graded potentials and spikes in the bipolar cell axon terminal to postsynaptic ganglion cells. Aim 2 will determine the influence of inhibitory feedback to bipolar cells on their spike-like events, using whole-cell patch clamp in retinal slices and whole-mounts. Integrating the results from Aims 1 and 2, Aim 3 will develop realistic computational models that will enable us to test novel hypotheses about local processing functions performed by spikes and graded potentials in bipolar cell terminals. We will develop computer models of identified types of bipolar cell with different axon terminal morphologies. This will allow us to determine the critical parameters for independence of signals in bipolar cell axon terminals. In particular, we will explore the role of inhibitory feedback from amacrine cells in modulating spikes and graded signals in bipolar cell axon terminals, to support signal independence between terminals and control their vesicle release with precise timing. Overall, the project will reveal how mammalian bipolar cells transform the graded potentials they receive from photoreceptors with high fidelity into spike-like transient events in their axon terminals, and how these signals interact with inhibitory feedback to modulate bipolar synaptic output. The proposed research will improve our understanding of the signal processing of the adult retina. As bipolar cells are critical targets for stimulation by visual prostheses and genetic approaches to restoring vision loss from a range of eye diseases, the knowledge gained here will guide the further development of such devices and treatments.